The Arabidopsis gene PLEIOTROPIC REGULATORY LOCUS 1 (PRL1) encodes a protein that has been identified as a member of a conserved WDR protein family in eukaryotes. Mutations of PRL1 result in hypersensitivity to sugars, cause a wide range of pleiotropic alterations in root, leaf and flower development and activate responses to abiotic and biotic stress stimuli by modifying the plant hormone homeostasis. In contrast to other eukaryotes, the genome of Arabidopsis encodes a PRL1 paralogue, PRL2, which shares a highly conserved C-terminal WDR domain with PRL1. However, PRL2 carries divergent N-terminal sequences when compared to PRL1. Nevertheless, the N-terminal domains of PRL1 and PRL2 show an exclusive evolutionary conservation in the plant kingdom. Studies in yeast and mammals revealed that PRL1 plays a pivotal role in spliceosome activation. However, the divergence of the PRL1 N-terminal region in Arabidopsis implies plant-specific functions in splicing. One aim of this work was to provide insights into the regulatory functions of PRL1. Affymetrix ATH1 arrays and Tiling arrays, together with RNA-Seq analysis, confirmed genome-wide alterations of the prl1 mutant. Moreover, the up-regulation of transposable elements, together with the up-regulation of other genes suggests that PRL1 is involved in the control of gene silencing. Furthermore, it was shown by using modified PRL1 constructs that PRL1 transcription and PRL1 stability are stress-dependent and that PRL1 does not interfere with the proteasome activity. Another aim was to assess the degree of functional complementarity of PRL1 and PRL2. Therefore, the PRL2 transcription was compared to the one of PRL1, showing that PRL2 is transcribed in most organs at a significant lower level. In flowers, PRL2 is expressed specifically in male reproductive tissues, pollen grains, endosperm and the zygote, whereas PRL1 is active in developing ovules and in the integuments of developing seeds. The prl2 mutations cause embryo lethality, whereas somatic prl2 mosaics show a prl1-like phenotype. PRL1 and PRL2 sub-functionalization was revealed by the replacement of the divergent N-terminal domains of PRL1 with PRL2. The N-terminal domain of PRL2 rescues the root phenotype of prl1 only partially. However, it complements the embryonic lethality caused by prl2. Moreover, the N-terminal domain of PRL1 in combination with the C-terminal domain of PRL2 complements the phenotype of prl1, but not the embryo lethal phenotype of prl2. In addition to the different expression observed for PRL1 and PRL2, the diversification of the N-terminal domains has contributed to the sub-functionalization of PRL1 and PRL2 in Arabidopsis. The results presented in this work also suggest that PRL1 and PRL2 perform a specific function in the control of gene silencing in Arabidopsis.